Mammalian multidrug-resistance proteins (MRPs)

Subfamily C of the human ABC (ATP-binding cassette) superfamily contains nine proteins that are often referred to as the MRPs (multidrug-resistance proteins). The 'short' MRP/ABCC transporters (MRP4, MRP5, MRP8 and ABCC12) have a typical ABC structure with four domains comprising two membrane-spanning domains (MSD1 and MSD2) each followed by a nucleotide-binding domain (NBD1 and NBD2). The 'long' MRP/ABCCs (MRP1, MRP2, MRP3, ABCC6 and MRP7) have five domains with the extra domain, MSD0, at the N-terminus. The proteins encoded by the ABCC6 and ABCC12 genes are not known to transport drugs and are therefore referred to as ABCC6 and ABCC12 (rather than MRP6 and MRP9) respectively. A large number of molecules are transported across the plasma membrane by the MRPs. Many are organic anions derived from exogenous sources such as conjugated drug metabolites. Others are endogenous metabolites such as the cysteinyl leukotrienes and prostaglandins which have important signalling functions in the cell. Some MRPs share a degree of overlap in substrate specificity (at least in vitro), but differences in transport kinetics are often substantial. In some cases, the in vivo substrates for some MRPs have been discovered aided by studies in gene-knockout mice. However, the molecules that are transported in vivo by others, including MRP5, MRP7, ABCC6 and ABCC12, still remain unknown. Important differences in the tissue distribution of the MRPs and their membrane localization (apical in contrast with basolateral) in polarized cells also exist. Together, these differences are responsible for the unique pharmacological and physiological functions of each of the nine ABCC transporters known as the MRPs.     

Sex differences in drug action: the role of multidrug-resistance proteins (MRPs)

The mechanisms of participation of multidrug-resistance proteins (MRPs) in sex difference pattern of drug efficiency and side effects have been analyzed. MRPs structure, their tissue and cellular localization, substrate specificity, and functions have been considered. Regulation of expression and activity of MRPs by endogenous metabolites, signal compounds, including sex hormones, as well as by drug agents have been represented. The role of nuclear receptors in the regulation of MRPs expression has been demonstrated. The data on sex differences in hepatic and renal MRPs expression and expression of nuclear receptors, participating in their induction, have been shown. The participation of MRPs in the formation of sex differences of drug pharmacokinetics has been discussed. Sex differences in representation and functional activity of MRPs in such excretory organs as liver and kidney was concluded would play the essential role in sex dependence of pharmacokinetics and efficiency of drug action.     

Multidrug resistance during cancer chemotherapy — biotechnological solutions to a clinical problem

Tumour cells can be resistant to a variety of chemotherapeutic drugs of different structure (multidrug resistance) by expressing a transmembrane pump (P-glycoprotein) on their cell surface. This situation can lead to a failure of cancer chemotherapy as the P-glycoprotein acts by actively pumping the drugs out of cells, thus lowering the intracellular concentration of the drug and, hence, its cytotoxic effectiveness. This review summarizes present and proposed approaches to preventing or circumventing the action of this drug-transporting protein.     

Multidrug resistance (MDR) genes in haematological malignancies

The emergence of drug resistant cells is one of the main obstacles for successful chemotherapeutic treatment of haematological malignancies. Most patients initially respond to chemotherapy at the time of first clinical admission, but often relapse and become refractory to further treatment not only to the drugs used in the first treatment but also to a variety of other drugs. Laboratory investigations have now provided a cellular basis for this clinical observation of multidrug resistance (MDR). Expression of a glycoprotein (referred to as P-glycoprotein) in the membrane of cells made resistantin vitro to naturally occurring anticancer agents like anthracyclines, Vinca alkaloids and epipodophyllotoxins, has been shown to be responsible for the so-called classical MDR phenotype. P-glycoprotein functions as an ATP-dependent, unidirectional drug efflux pump with a broad substrate specificity, that effectively maintains the intracellular cytotoxic drug concentrations under a non-cytotoxic threshold value. Extensive clinical studies have shown that P-glycoprotein is expressed on virtually all types of haematological malignancies, including acute and chronic leukaemias, multiple myelomas and malignant lymphomas. Since in model systems for P-glycoprotein-mediated MDR, drug resistance may be circumvented by the addition of non-cytotoxic agents that can inhibit the outward drug pump, clinical trials have been initiated to determine if such an approach will be feasible in a clinical situation. Preliminary results suggest that some haematological malignancies, among which are acute myelocytic leukaemia, multiple myeloma and non-Hodgkin's lymphoma, might benefit from the simultaneous administration of cytotoxic drugs and P-glycoprotein inhibitors. However, randomised clinical trials are needed to evaluate the use of such resistance modifiers in the clinic.Abbreviations ALL acute lymphocytic leukaemia - AML acute myelocytic leukaemia - BM bone marrow - CAT chloramphenicol acetyltransferase - CLL chronic lymphocytic leukaemia - CML chronic myelocytic leukaemia - CR complete remission - HCL hairy cell leukaemia - MDR multidrug resistance - MDS myelodysplastic syndrome - MM multiple myeloma - MoAb monoclonal antibody - NHL non-Hodgkin's lymphoma - PB peripheral blood - PCR polymerase chain reaction - PLL prolymphocytic leukaemia - RMA resistance modifying agent - VAD vincristine, doxorubicin, dexamethasone     

Multidrug transporter ABCG2/breast cancer resistance protein secretes riboflavin (vitamin B2) into milk

The multidrug transporter breast cancer resistance protein (BCRP/ABCG2) is strongly induced in the mammary gland during pregnancy and lactation. We here demonstrate that BCRP is responsible for pumping riboflavin (vitamin B(2)) into milk, thus supplying the young with this important nutrient. In Bcrp1(-/-) mice, milk secretion of riboflavin was reduced >60-fold compared to that in wild-type mice. Yet, under laboratory conditions, Bcrp1(-/-) pups showed no riboflavin deficiency due to concomitant milk secretion of its cofactor flavin adenine dinucleotide, which was not affected. Thus, two independent secretion mechanisms supply vitamin B(2) equivalents to milk. BCRP is the first active riboflavin efflux transporter identified in mammals and the first transporter shown to concentrate a vitamin into milk. BCRP activity elsewhere in the body protects against xenotoxins by reducing their absorption and mediating their excretion. Indeed, Bcrp1 activity increased excretion of riboflavin into the intestine and decreased its systemic availability in adult mice. Surprisingly, the paradoxical dual utilization of BCRP as a xenotoxin and a riboflavin pump is evolutionarily conserved among mammals as diverse as mice and humans. This study establishes the principle that an ABC transporter can transport a vitamin into milk and raises the possibility that other vitamins and nutrients are likewise secreted into milk by ABC transporters.     

Multidrug resistance and behavioural phenotype of cancer cells

Resistance to cytotoxic chemotherapy is a major obstacle preventing successful treatment of cancer, allowing dissemination of tumour metastases, and may be viewed as the ultimate cause of death in the majority of patients with a malignant disease. Although cytotoxic chemotherapy is classically employed to produce maximal killing of malignant cells, the therapeutic doses of individual drugs required to achieve this objective are, in general, highly toxic to non-neoplastic host tissues. However, there are several different aspects of cancer cell biology, distinct from their susceptibility to cytotoxicity, that might be exploited in order to alter the behavioral phenotypes of malignant neoplasms. Such features include regulation of cell proliferation, tumorigenicity and metastatic potential Non-cytotoxic modulation of malignant cells may provide an alternative, and more effective, method of controlling the aggressive behaviour of cancer cells while exhibiting less iatrogenic morbidity and mortality than the therapeutic regimens presently employed.     

Multidrug resistance protein 4 (MRP4/ABCC4) mediates efflux of bimane-glutathione

Multidrug resistance proteins (MRPs) are ATP-dependent export pumps that mediate the export of organic anions. ABCC1 (MRP1), ABCC2 (MRP2) and ABCC3 (MRP3) are all able to facilitate the efflux of anionic conjugates including glutathione (GSH), glucuronide and sulfate conjugates of xenobiotics and endogenous molecules. Earlier studies showed that ABCC4 functions as an ATP-driven export pump for cyclic AMP and cyclic GMP, as well as estradiol-17-beta-D-glucuronide. However, it was unclear if other conjugated metabolites can be transported by ABCC4. Hence in this study, a fluorescent substrate, bimane-glutathione (bimane-GS) was used to further examine the transport activity of ABCC4. Using cells stably overexpressing ABCC4, this study shows that ABCC4 can facilitate the efflux of the glutathione conjugate, bimane-glutathione. Bimane-glutathione efflux increased with time and >85% of the conjugate was exported after 15min. This transport was abolished in the presence of 2.5microM carbonylcyanide m-chlorophenylhydrasone (CCCP), an uncoupler of oxidative phosphorylation. Inhibition was also observed with known inhibitors of MRP transporters including benzbromarone, verapamil and indomethacin. In addition, 100microM methotrexate, an ABCC4 substrate or 100microM 6-thioguanine (6-TG), a compound whose monophosphate metabolite is an ABCC4 substrate, reduced efflux by >40%. A concentration-dependent inhibition of bimane-glutathione efflux was observed with 1-chloro-2,4-dinitrobenzene (CDNB) which is metabolized intracellularly to the glutathione conjugate, 2,4-dinitrophenyl-glutathione (DNP-GS). The determination that ABCC4 can mediate the transport of glucuronide and glutathione conjugates indicates that ABCC4 may play a role in the cellular extrusion of Phase II detoxification metabolites.     

Multidrug resistance in fungi

Antibiotic and synthetic chemotherapeutic resistance in pathogenic yeast becomes one of the biggest challenges for the modern chemotherapy. An increasing number of pathogenic yeast and filamentous fungi resistant to the action of the majority of currently used drugs is isolated in clinics nowadays. Among variety of the resistance mechanisms, the most dangerous grows to be the multidrug resistance. The most important mechanism of the multidrug resistance is the overexpression of membrane proteins participating in the active efflux of drugs out of the cells subjected to chemotherapy. Representatives of two classes of multidrug efflux transporters, ABC and MFS, have been identified in fungi. One of the most important strategies for overcome the phenomenon of multidrug resistance in pathogenic fungi, is the use of chemical compounds co-administrated with chemotherapeutics which are able to restore drug susceptibility in multidrug resistant cells. Mode of action of these chemical compounds may be very diverse, from the substrate competition, through the influence on the membrane fluidity, to the multidrug transporters activity modulation. This paper presents a review of the current knowledge on proteins contributing to fungal multidrug resistance and strategies for overcoming multidrug resistance by pharmacological intervention.     

Multidrug chemotherapy of osteogenic sarcoma (author's transl)]

Osteogenic sarcoma may be treated effectively by radical surgical removal of the primary tumor and combined chemotherapy, including Adriamycin and high dose Methotrexate. In order to render any protocol a safe procedure, strict precautions are required to avoid drug toxicity. We present a protocol, "COSS 77", presently employed in several university hospitals of West Germany and Austria. Final results concerning long term prognosis and long term side effects are not yet available.     

Identification of mammalian mitochondrial ribosomal proteins (MRPs) by N-terminal sequencing of purified bovine MRPs and comparison to data bank sequences: the large subribosomal particle

Bovine mitochondrial ribosomes are presented as a model system for mammalian mitochondrial ribosomes. An alternative system for identifying individual bovine mitochondrial ribosomal proteins (MRPs) by RP-HPLC is described. To identify and to characterize individual MRPs proteins were purified from bovine liver, separated by RP-HPLC, and identified by 2D PAGE techniques and immunoblotting. Molecular masses of individual MRPs were determined. Selected proteins were subjected to N-terminal amino acid sequencing. The peptide sequences obtained were used to screen different databases to identify several corresponding MRP sequences from human, mouse, rat, and yeast. Signal sequences for mitochondrial import were postulated by comparison of the bovine mature N-termini determined by amino acid sequencing with the deduced mammalian MRP sequences. Significant sequence similarities of these new MRPs to known r-proteins from other sources, e.g., E. coli, were detected only for two of the four MRP families presented. This finding suggests that mammalian mitochondrial ribosomes contain several novel proteins. Amino acid sequence information for all of the bovine MRPs will prove invaluable for assigning functions to their genes, which would otherwise remain unknown.     

多药耐药基因与子宫颈癌化疗

肿瘤细胞对化疗药物产生多药耐药性是化疗失败的主要原因之一,肿瘤多药耐药的机制十分广泛,其中P-gp/mdrl介导的多药耐药是最经典的耐药机制.故本文就MDR1与宫颈癌化疗的关系进行回顾和总结.     

Multidrug chemotherapy of tuberculosis in rhesus monkeys

Occurrence of tuberculosis caused by Mycobacterium bovis in a colony of rhesus monkeys allowed evaluation of a modern multidrug therapeutic regimen. Fifteen tuberculin positive rhesus monkeys with disseminated tuberculosis were evaluated for extent of disease by radiographic techniques, physical examination and laparotomy prior to treatment. Monkeys were divided into treatment groups of 3, 6 and 12 months duration and were treated once daily with isoniazid, rifampin and ethambutol. All animals survived their treatment course, had marked clinical improvement and rapid resolution of radiographically demonstrable lesions. Lesion regression evaluated by necropsy and histopathology correlated positively with length of treatment interval. Mycobacterium bovis was not isolated from any animal following treatment. Multidrug chemotherapy of tuberculosis was considered successful and practical in rhesus monkeys at the 12 month treatment interval. Chemotherapy may provide a reasonable alternative to destruction of valuable animals infected with tuberculosis.     

Multidrug resistance phosphoglycoprotein (ABCB1) in the mouse placenta: fetal protection

The multidrug resistance phosphoglycoprotein ATP-binding cassette subfamily B (ABCB1) actively extrudes a range of structurally and functionally diverse xenobiotics as well as glucocorticoids. ABCB1 is present in many cancer cell types as well as in normal tissues. Although it has been localized within the mouse placenta, virtually nothing is known about its regulation. In the mouse, two genes, Abcb1a and Abcb1b, encode ABCB1. We hypothesized that there are changes in placental Abcb1a and Abcb1b gene expression and ABCB1 protein levels during pregnancy. Using in situ hybridization, we demonstrated that Abcb1b mRNA is the predominant placental isoform and that there are profound gestational changes in the expression of both Abcb1a and Abcb1b mRNA. Placentas from pregnant mice were analyzed between Embryonic Days (E) 9.5 and 19 (term approximately 19.5d). Abcb1b mRNA was detected in invading trophoblast cells by E9.5, peaked within the placental labyrinth at E12.5, and then progressively decreased toward term (P < 0.0001). Abcb1a mRNA, although lower than that of Abcb1b at midgestation, paralleled changes in Abcb1b mRNA. Changes in Abcb1 mRNA were reflected by a significant decrease in ABCB1 protein (P < 0.05). A strong correlation existed between placental Abcb1b mRNA and maternal progesterone concentrations, indicating a potential role of progesterone in regulation of placental Abcb1b mRNA. In conclusion, there are dramatic decreases in Abcb1a and Abcb1b mRNA and in ABCB1 at the maternal-fetal interface over the second half of gestation, suggesting that the fetus may become increasingly susceptible to the influences of xenobiotics and natural steroids in the maternal circulation.     

Multidrug resistance evaluation by confocal microscopy in primary urothelial cancer explant colonies

Assessing functional multidrug resistance (MDR) status in clinical biopsy material using drug autofluorescence has potential applications to clinical management. The small size of many cystoscopy specimens has led us to develop, as an alternative to flow cytometry, a protocol for studying epirubicin accumulation in adherent colonies of primary bladder cancer cells viewed live andin situ by confocal microscopy. The limitations to quantitation inherent in this technique are compensated for by preservation of cellular organisation and the elimination of non-malignant cells. Biopsy material is disaggregated and explanted into culture-grade petri dishes. After incubation for three to seven days plaques of epithelial cells have developed. Classical patterns of sensitive and resistant drug distribution are observed. Cells of the rolled edges of the colony accumulate more drug than those of the inner epithelial monolayer. Some central areas of larger colonies give the appearance of drug arrested at the intercellular junctions to give a fenestrated pattern. These observations contribute to the understanding of mechanisms in MDR as well as forming the basis for a clinical urological MDR evaluation protocol.     

Multidrug resistance (MDR1) P-glycoprotein enhances esterification of plasma membrane cholesterol

Class I P-glycoproteins (Pgp) confer multidrug resistance in tumors, but the physiologic function of Pgp in normal tissues remains uncertain. In cells derived from tissues that normally express Pgp, recent data suggest a possible role for Pgp in cholesterol trafficking from the plasma membrane to the endoplasmic reticulum. We investigated the esterification of plasma membrane cholesterol under basal conditions and in response to sphingomyelinase treatment in transfected and drug-selected cell lines expressing differing amounts of functional class I Pgp. Compared with parental NIH 3T3 fibroblasts, cells transfected with human multidrug resistance (MDR1) Pgp esterified more cholesterol both without and with sphingomyelinase. Esterification also was greater in drug-selected Dox 6 myeloma cells than parental 8226 cells, which express low and non-immunodetectable amounts of Pgp, respectively. However, no differences in total plasma membrane cholesterol were detected. Transfection of fibroblasts with the multidrug resistance-associated protein (MRP) did not alter esterification, showing that cholesterol trafficking was not generally affected by ATP-binding cassette transporters. Steroidal (progesterone, dehydroepiandrosterone) and non-steroidal antagonists (verapamil, PSC 833, LY335979, and GF120918) were evaluated for effects on both cholesterol trafficking and the net content of 99mTc-Sestamibi, a reporter of drug transport activity mediated by Pgp. In Pgp-expressing cells treated with nonselective and selective inhibitors, both the kinetics and efficacy of inhibition of cholesterol esterification differed from the antagonism of drug transport mediated by Pgp. Thus, although the data show that greater expression of class I Pgp within a given cell type is associated with enhanced esterification of plasma membrane cholesterol in support of a physiologic function for Pgp in facilitating cholesterol trafficking, the molecular mechanism is dissociated from the conventional drug transport activity of Pgp.